Nitric oxide (NO), carbon monoxide (CO), and molecular oxygen (O2) are three key gaseous messengers in mammals. Although significant advances have been made in elucidating the mechanisms by which these molecules are biosynthesized, very little is known about how they transduce signals to trigger a physiological response. NO mediates its physiological actions via its heme protein receptor, soluble guanylyl cyclase (sGC). Despite three decades of research, the structural basis of how NO binding to sGC activates second messenger (cGMP) production is not understood. We have devised innovative strategies to gain insights into this process. This includes our discovery of a novel family of bacterial NO sensors (SONO) that share remarkable sequence identity with the sensor domain of sGC. Although the signal for SONO has remained unchanged during two billion years of evolution, its function is unrelated to cGMP biosynthesis. The SONO-protein interactions we have identified has allowed us to frame the provocative hypothesis that the NO sensor is a promiscuous molecular switch capable of triggering functionally unrelated signaling events via a limited set of structural changes. Four specific aims have been designed to test this hypothesis: Aim 1. What is the structure of NO- and CO-activated SONO? Aim 2: What are the structural bases of heme- and NO-independent signaling by human sGC? Aim 3: How is NO and activator binding coupled to sGC catalysis? Aim 4: What is the molecular mechanism by which SONO mediates chemotaxis in human pathogens? The proposed research will provide novel insights into mechanisms of gaseous messenger signaling. It will also pave the way for rational strategies aimed at developing therapeutic interventions for cardiovascular diseases. PUBLIC HEALTH RELEVANCE: Among the several functions of nitric oxide - a gas produced by the cells lining our blood vessels - is its ability to convert sexual excitement into erections in males. This proposal is aimed at understanding how nitric oxide works in healthy and disease states. [unreadable] [unreadable] [unreadable]